The level of xanthine oxidase (“XO”) in an animal increases markedly (>400-fold in bronchoalveolar fluid in pneumonitis) during inflammation, ischemia-reperfusion injury, and atherosclerosis. Particularly, due to the spillover of tissue XO into the circulation, plasma levels of XO may be detected in an animal experiencing adult respiratory distress syndrome, ischemia-reperfusion injury, arthritis, sepsis, hemorrhagic shock, and other inflammatory conditions. Inflammatory-induced histamine release by mast cells and basophils also enhances the activity of XO.
Superoxide radical (O2−) can be generated by xanthine oxidase and NADPH oxidase from the partial reduction of molecular oxygen. Neutrophils and macrophages are known to produce O2− and hydrogen peroxide (H2O2), which normally are involved in the killing of ingested or invading microbes (T. Oda et al., Science, 244:974–976). Under physiologic conditions XO is ubiquitously present in the form of a xanthine dehydrogenase (XDH). XDH is a molybdenum iron-sulfur flavin dehydrogenase that uses NAD+ as an electron acceptor to oxidize purines, pyrimidines, pteridins, and other heterocyclic nitrogen-containing compounds. In mammals, XDH is converted from the NAD-dependent dehydrogenase form to the oxygen-dependent oxidase form, either by reversible sulfhydryl oxidation or irreversible proteolytic modification (S. Tan et al., Free Radic. Biol. Med. 15:407–414). Xanthine oxidase then no longer uses NAD+ as an electron acceptor, but transfers electrons onto oxygen, generating O2−, H2O2, and hydroxyl radical (OH) as purines are degraded to uric acid (J. M. McCord et al., New Engl. J. Med. 312:159–163; R. Miesel et al., Inflammation, 18:597–612). Inflammatory activation converts XDH to XO, mainly by oxidizing structurally important thiolates. Inflammation also markedly up-regulates the conversion of xanthine dehydrogenase (T. D. Engerson et al., J. Clin. Invest. 79:1564–1570).
Inhibition of XO activity blocks the formation of O2− and prevents loss of purine nucleotides, and is therefore salutary in a variety of shock and ischemia reperfusion disorders. Pharmacologic inhibition of XO can also be beneficial by blocking the pro-inflammatory effect of O2− on gene expression (M. D. Schwartz et al., Am. J. Respir. Cell. Mol. Biol., 12:434–440). O2− has been implicated in the nuclear translocation of NF-kappa B and the expression of NF-icB-dependent genes. In mice subjected to hemorrhagic shock, depletion of XO by a tungsten-enriched diet decreased mononuclear mRNA levels of IL-113 and TNF-a. Similar results were obtained after pharmacologic inhibition of XO by in vivo administration of allopurinol. A vicious cycle can be created by oxidant stress, in which O2− induction of pro-inflammatory cytokines results in greater XDH to XO conversion, and thus more O2− production. This suggests that XO inhibitors can exert important anti-inflammatory actions by interrupting this process at multiple points, in particular, by blocking pro-inflammatory gene expression.
Pharmacologic inhibition of XO can also be beneficial in hemorrhagic shock by preserving the intracellular nucleotide pool. Under conditions of energetic failure, induced by hypoxia or by oxidant-induced poly(ADP-ribose) synthetase activation, high-energy phosphate nucleotides are sequentially degraded to inosine monophosphate→xanthine→hypoxanthine. In the presence of XO and molecular oxygen, xanthine and hypoxanthine degrade to uric acid, thereby depleting the purine pool. The loss of available purines with which to form ATP accelerates the loss of intracellular energetics and contributes to cell necrosis and organ failure. XO inhibitors block this terminal degradative pathway and permit the cell to recover and reestablish adequate stores of high energy phosphate nucleotides. In a canine model of severe hemorrhagic shock, pre-treatment with allopurinol resulted in a 6-fold increase in survival (J. W. Crowell et al., Am. J. Phys. 216:744–748). When the administration of allopurinol was delayed until after shock had been produced, allopurinol exerted no benefit. Infusion of the purine base hypoxanthine after the onset of shock similarly provided no benefit. When allopurinol and hypoxanthine were co-infused, however, there was a dramatic increase in survival (no survival in control group at 16 hours post-shock vs. a 40% survival in the treated group at 48 hours). Similar results were obtained in a canine model of hemorrhagic shock in which allopurinol significantly improved survival, whereas a cocktail of free-radical scavengers (superoxide dismutase, catalase, dimethylsulfoxide, and alpha tocopherol) had no effect (D. Mannion, et al., Circ. Shock, 42:39–43). Thus, XO blockade appears to be beneficial by three independent mechanisms: blockade of O2− formation; inhibition of O2− mediated pro-inflammatory gene expression; and preservation of the nucleotide pool available for ATP formation.
Accordingly, there is a clear need for compounds that inhibit the levels of xanthine oxidase in an animal and, accordingly, that are useful for treating or preventing an inflammation disease, a reperfusion disease, hyperuricemia, gout, tumor-lysis syndrome, or an inflammatory bowel disorder.
Citation of any reference in Section 2 of this application is not an admission that the reference is prior art to the application.